AbstractOrganisms are classified into three domains, Prokaryota, Archaea, and Eukaryota, and their evolutionary divergence has been characterized based on morphological and molecular features using rationale based on Darwin’s theory of natural selection. However, universal rules that govern genome evolution have not been identified. Here, a simple, innovative approach has been developed to evaluate biological evolution initiating the origin of life: whole genomes were divided into several fragments, and then differences in normalized nucleotide content between nucleotide pairs were compared. Based on nucleotide content structures, Monosiga brevicollis mitochondria may be the most primitive extant ancestor of the species examined here. The two normalized nucleotide contents are universally expressed by a linear regression line, (X − Y)/(X + Y) = a (X − Y) + b, where X and Y are nucleotide contents and (a) and (b) are constants. The value of (G + C), (G + A), (G + T), (C + A), (C + T) and (A + T) was ~0.5. Plotting (X − Y)/(X + Y) against X/Y showed a logarithmic function (X − Y)/(X + Y) = a ln X/Y + b, where (a) and (b) are constant. Nucleotide content changes are expressed by a definitive equation, (X − Y) ≈ 0.25 ln(X/Y).